Disruptions in energy supply and utilization within the brain have severe functional consequences. In particular, the loss of the creatine (Cr)-phosphocreatine (PCr) shuttle leads to severe intellectual disability, epilepsy and aphasia. The Cr-PCr shuttle is responsible for rapid ATP replenishment. The most prevalent cause of Cr loss in the brain is due to mutations in the X-linked Cr transporter (CrT). To date, there are no treatments available for CrT deficiency. In order to better understand the pathophysiology of CrT deficiency as well as to develop and screen treatments, we generated CrT knockout (CrT-/y) mice that show severe cognitive deficits similar to CrT deficient patients. While Cr deficiency is the root of the cognitive disorders, it likely does not play a direct role in cognitive function. I is likely that the loss of Cr leads to larger disruptions of systems directly involved in neuronal function. It is essential to identify these systems and determine how the loss of Cr affects their function. Na+,K +-ATPases (NKA) are essential for proper cognitive function and consume 50-60% of the brain's resting metabolic output. Previous studies, along with preliminary data from our lab, show that Cr is essential for proper function of NKA. The mechanisms underlying the role of the Cr-PCr shuttle on NKA function has not been elucidated. The purpose of this proposal is to identify the mechanisms underlying the interaction between NKA and Cr. The hypothesis for this proposal is that the rapid ATP turnover provided by Cr-PCr shuttle is required for proper NKA activity. In aim 1, the role of each component of the Cr-PCr shuttle (Cr, PCr, ATP) on NKA function will be evaluated. The effects of a potential treatment for Cr deficiency, cyclocreatine, on NKA function will be determined as well. In aim 2, the effect of Cr-deficiency on the individual NKA catalytic subunits function will be evaluated. Each subunit has a unique function within the brain, making it essential to determine how the loss of Cr disrupts their function. Upon the completion of these studies, a direct link between the loss of Cr and NKA function will be established, shedding light onto the function of NKA, an essential protein for brain function as well as advancing the understanding of CrT deficiency, a significant human disorder.